Arrhythmia Score Research Articles

Gabapentin alleviates myocardial ischemia–reperfusion injury by increasing the protein expression of GABAARδ

Gabapentin is a commonly used analgesic in the clinic to reduce opioid consumption. It is well known that gabapentin can reduce cerebral ischemia-reperfusion injury (IRI). However, it remains unclear whether gabapentin can reduce myocardial IRI. Before the performance of myocardial ischemia and reperfusion (I/R), rats received gabapentin without or with an intravenous injection of PI3K inhibitor (LY294002), or an intraspinal injection of lentivirus-mediated GABAARδ-shRNA. The myocardial IRI were evaluated by calculating the infarction area, arrhythmia score and myocardial apoptosis. The activity of PI3K/Akt and the expression of GABAARδ were quantified by western blotting. The effect of gabapentin on myocardial I/R was further demonstrated in vitro by establishing oxygen-glucose deprivation and reoxygenation in cardiomyocytes. After I/R in vivo, there were significant increases in infarction area, arrhythmia and Bax protein expression in the myocardium, as well as a decrease of GABAARδ in the spinal cord. Meanwhile, I/R also decreased the protein expression of PI3K/Akt and Bcl-2. Gabapentin pretreatment successfully attenuated IRI including reducing the myocardial infarction area and apoptosis. This effect was abolished by both the systemic inhibition of PI3K/Akt and the intraspinal suppression of GABAARδ. However, gabapentin pretreatment failed to prevent cellular injury induced by OGD/R in cardiomyocytes. Therefore, the myocardial protective effect of gabapentin may be attributed to activating PI3K/Akt in the myocardium and upregulating GABAARδ in the spinal cord. Gabapentin achieved a potent protective effect on the myocardium during the course of routine clinical treatment.

GABAergic Signaling during Spinal Cord Stimulation Reduces Cardiac Arrhythmias in a Porcine Model.

Neuraxial modulation, including spinal cord stimulation, reduces cardiac sympathoexcitation and ventricular arrhythmogenesis. There is an incomplete understanding of the molecular mechanisms through which spinal cord stimulation modulates cardiospinal neural pathways. The authors hypothesize that spinal cord stimulation reduces myocardial ischemia-reperfusion-induced sympathetic excitation and ventricular arrhythmias through γ-aminobutyric acid (GABA)-mediated pathways in the thoracic spinal cord. Yorkshire pigs were randomized to control (n = 11), ischemia-reperfusion (n = 16), ischemia-reperfusion plus spinal cord stimulation (n = 17), ischemia-reperfusion plus spinal cord stimulation plus γ-aminobutyric acid type A (GABAA) or γ-aminobutyric acid type B (GABAB) receptor antagonist (GABAA, n = 8; GABAB, n = 8), and ischemia-reperfusion plus GABA transaminase inhibitor (GABAculine, n = 8). A four-pole spinal cord stimulation lead was placed epidurally (T1 to T4). GABA modulating pharmacologic agents were administered intrathecally. Spinal cord stimulation at 50 Hz was applied 30 min before ischemia. A 56-electrode epicardial mesh was used for high-resolution electrophysiologic recordings, including activation recovery intervals and ventricular arrhythmia scores. Immunohistochemistry and Western blots were performed to measure GABA receptor expression in the thoracic spinal cord. Cardiac ischemia led to myocardial sympathoexcitation with reduction in activation recovery interval (mean ± SD, -42 ± 11%), which was attenuated by spinal cord stimulation (-21 ± 17%, P = 0.001). GABAA and GABAB receptor antagonists abolished spinal cord stimulation attenuation of sympathoexcitation (GABAA, -9.7 ± 9.7%, P = 0.043 vs. ischemia-reperfusion plus spinal cord stimulation; GABAB, -13 ± 14%, P = 0.012 vs. ischemia-reperfusion plus spinal cord stimulation), while GABAculine alone caused a therapeutic effect similar to spinal cord stimulation (-4.1 ± 3.7%, P = 0.038 vs. ischemia-reperfusion). The ventricular arrhythmia score supported these findings. Spinal cord stimulation during ischemia-reperfusion increased GABAA receptor expression with no change in GABAB receptor expression. Thoracic spinal cord stimulation reduces ischemia-reperfusion-induced sympathoexcitation and ventricular arrhythmias through activation of GABA signaling pathways. These data support the hypothesis that spinal cord stimulation-induced release of GABA activates inhibitory interneurons to decrease primary afferent signaling from superficial dorsal horn to sympathetic output neurons in the intermediolateral nucleus.

Thoracic dorsal root ganglion stimulation reduces acute myocardial ischemia induced ventricular arrhythmias.

Dorsal root ganglion stimulation (DRGS) may serve as a novel neuromodulation strategy to reduce cardiac sympathoexcitation and ventricular excitability. In this pre-clinical study, we investigated the effectiveness of DRGS on reducing ventricular arrhythmias and modulating cardiac sympathetic hyperactivity caused by myocardial ischemia. Twenty-three Yorkshire pigs were randomized to two groups, which was control LAD ischemia-reperfusion (CONTROL) or LAD ischemia-reperfusion + DRGS (DRGS) group. In the DRGS group (n = 10), high-frequency stimulation (1 kHz) at the second thoracic level (T2) was initiated 30 min before ischemia and continued throughout 1 h of ischemia and 2 h of reperfusion. Cardiac electrophysiological mapping and Ventricular Arrhythmia Score (VAS) were assessed, along with evaluation of cFos expression and apoptosis in the T2 spinal cord and DRG. DRGS decreased the magnitude of activation recovery interval (ARI) shortening in the ischemic region (CONTROL: -201 ± 9.8 ms, DRGS: -170 ± 9.4 ms, p = 0.0373) and decreased global dispersion of repolarization (DOR) at 30 min of myocardial ischemia (CONTROL: 9546 ± 763 ms2, DRGS: 6491 ± 636 ms2, p = 0.0076). DRGS also decreased ventricular arrhythmias (VAS-CONTROL: 8.9 ± 1.1, DRGS: 6.3 ± 1.0, p = 0.038). Immunohistochemistry studies showed that DRGS decreased % cFos with NeuN expression in the T2 spinal cord (p = 0.048) and the number of apoptotic cells in the DRG (p = 0.0084). DRGS reduced the burden of myocardial ischemia-induced cardiac sympathoexcitation and has a potential to be a novel treatment option to reduce arrhythmogenesis.

Effect of total flavonoids of buckwheat flower and leaf on myocardial cell apoptosis and Wnt/β-catenin/PPARγ pathway in arrhythmic rats

This paper aimed to investigate the effect of total flavonoids of buckwheat flower and leaf on myocardial cell apoptosis and Wnt/β-catenin/peroxisome proliferator-activated receptor γ(PPARγ) pathway in arrhythmic rats. SD rats were randomly divided into a control group, a model group, a low-dose(20 mg·kg~(-1)) group of total flavonoids of buckwheat flower and leaf, a medium-dose(40 mg·kg~(-1)) group of total flavonoids of buckwheat flower and leaf, a high-dose(80 mg·kg~(-1)) group of total flavonoids of buckwheat flower and leaf, a propranolol hydrochloride(2 mg·kg~(-1)) group, with 12 rats in each group. Except the control group, rats in other groups were prepared as models of arrhythmia by sublingual injection of 1 mL·kg~(-1) of 0.002% aconitine. After grouping and intervention with drugs, the arrhythmia, myocardial cells apoptosis, myocardial tissue glutathione peroxidase(GSH-Px), catalase(CAT), malondialdehyde(MDA), serum interleukin-6(IL-6), prostaglandin E2(PGE2) levels, myocardial tissue apoptosis, and Wnt/β-catenin/PPARγ pathway-related protein expression of rats in each group were measured. As compared with the control group, the arrhythmia score, the number of ventricular premature beats, ventricular fibrillation duration, myocardial cell apoptosis rate, MDA levels in myocardial tissues, serum IL-6 and PGE2 levels, Bax in myocardial tissues, and Wnt1 and β-catenin protein expression levels increased significantly in the model group, whereas the GSH-Px and CAT levels, and Bcl-2 and PPARγ protein expression levels in myocardial tissues reduced significantly. As compared with the model group, the arrhythmia score, the number of ventricular premature beats, ventricular fibrillation duration, myocardial cell apoptosis rate, MDA leve in myocardial tissues, serum IL-6 and PGE2 levels, Bax in myocardial tissues, and Wnt1 and β-catenin protein expression levels reduced in the drug intervention groups, whereas the GSH-Px and CAT levels and Bcl-2 and PPARγ protein expression levels in myocardial tissues increased. The groups of total flavonoids of buckwheat flower and leaf were in a dose-dependent manner. There was no significant difference in the levels of each index in rats between the propranolol hydrochloride group and the high-dose group of total flavonoids of buckwheat flower and leaf. The total flavonoids of buckwheat flower and leaf inhibit the activation of Wnt/β-catenin pathway, up-regulate the expression of PPARγ, reduce oxidative stress and inflammatory damage in myocardial tissues of arrhythmic rats, reduce myocardial cell apoptosis, and improve the symptoms of arrhythmia in rats.

Influence of verapamil on pressure overload-induced ventricular arrhythmias by regulating gene-expression profiles.

Life-threatening ventricular arrhythmias can lead to sudden cardiac death in patients. This study aimed to investigate the changes in gene profiles involved when verapamil (VRP) affects increased wall stress (pressure overload)-induced ventricular arrhythmias, thus revealing the potential causative molecular mechanisms and therapeutic targets through gene-expression identification and functional analysis. Animal models with wall stress-induced ventricular arrhythmias were established. Low (0.5 mg/kg) and high (1 mg/kg) doses of VRP were administered intravenously 10 minutes before transverse aortic constriction, and average ventricular arrhythmia scores were calculated. Next, we evaluated the molecular role of VRP by characterising differential gene-expression profiles between VRP-pretreated (1 mg/kg) and control groups using RNA-sequencing technology. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were used to reveal molecular function. A protein-protein interaction (PPI) network was then developed. VRP exerted its anti-arrhythmic effects in response to increases in left ventricular (LV) afterload. We detected differentially expressed genes (DEGs), of which 36 were upregulated and 1 397 downregulated, between the VRP-pretreated and model groups during acute increases in LV wall stress. GO analysis demonstrated that the DEGs were associated with cytoskeletal protein binding. KEGG analysis showed that enriched pathways were mainly distributed in adherens junctions, actin cytoskeleton regulation and the MAPK signalling pathway. Centralities analysis of the PPI identified Rac1, Grb2, Rbm8a and Mapk1 as hub genes. VRP prevented acute pressure overload-induced ventricular arrhythmias, possibly through the hub genes Rac1, Grb2, Rbm8a and Mapk1 as potential targets of VRP.

Repeatability of ventricular arrhythmia characteristics on the exercise-stress test in RYR2-mediated catecholaminergic polymorphic ventricular tachycardia.

In catecholaminergic polymorphic ventricular tachycardia (CPVT), the exercise-stress test (EST) is the cornerstone for the diagnosis, risk stratification, and assessment of therapeutic efficacy, but its repeatability is unknown. We aimed to test the repeatability of ventricular arrhythmia characteristics on the EST in patients with CPVT. EST-pairs (ESTs performed within 18 months between 2005 and 2021, on the same protocol, and without or on the exact same treatment) of patients with RYR2-mediated CPVT from two specialized centres were included. The primary endpoint was the repeatability of the maximum ventricular arrhythmia score [VAS: 0 for the absence of premature ventricular contractions (PVCs); 1 for isolated PVCs; 2 for bigeminal PVCs; 3 for couplets; and 4 for non-sustained ventricular tachycardia]. Secondary outcomes were the repeatability of the heart rate at the first PVC and the ΔVAS (the absolute difference in VAS between the EST-pairs). A total of 104 patients with 349 EST-pairs were included. The median duration between ESTs was 343 (interquartile range, 189-378) days. Sixty (17.2%) EST-pairs were off therapy. The repeatability of the VAS was moderate {Krippendorf α, 0.56 [95% confidence interval (CI), 0.48-0.64]}, and the repeatability of the heart rate at the first PVC was substantial [intra-class correlation coefficient, 0.78 (95% CI, 0.71-0.84)]. The use of medication was associated with a higher odds for a ΔVAS > 1 (odds ratio = 3.52; 95% CI, 2.46-4.57; P = 0.020). The repeatability of ventricular arrhythmia characteristics was moderate to substantial. This underlines the need for multiple ESTs in CPVT patients and CPVT suspicious patients and it provides the framework for assessing the therapeutic efficacy of novel CPVT therapies.

Spinal cord astrocytes regulate myocardial ischemia-reperfusion injury.

Astrocytes play a key role in the response to injury and noxious stimuli, but its role in myocardial ischemia-reperfusion (I/R) injury remains largely unknown. Here we determined whether manipulation of spinal astrocyte activity affected myocardial I/R injury and the underlying mechanisms. By ligating the left coronary artery to establish an in vivo I/R rat model, we observed a 1.7-fold rise in glial fibrillary acidic protein (GFAP) protein level in spinal cord following myocardial I/R injury. Inhibition of spinal astrocytes by intrathecal injection of fluoro-citrate, an astrocyte inhibitor, decreased GFAP immunostaining and reduced infarct size by 29% relative to the I/R group. Using a Designer Receptor Exclusively Activated by Designer Drugs (DREADD) chemogenetic approach, we bi-directionally manipulated astrocyte activity employing GFAP promoter-driven Gq- or Gi-coupled signaling. The Gq-DREADD-mediated activation of spinal astrocytes caused transient receptor potential vanilloid 1 (TRPV1) activation and neuropeptide release leading to a 1.3-fold increase in infarct size, 1.2-fold rise in serum norepinephrine level and higher arrhythmia score relative to I/R group. In contrast, Gi-DREADD-mediated inhibition of spinal astrocytes suppressed TRPV1-mediated nociceptive signaling, resulting in 35% reduction of infarct size and 51% reduction of arrhythmia score from I/R group, as well as lowering serum norepinephrine level from 3158 ± 108 to 2047 ± 95pg/mL. Further, intrathecal administration of TRPV1 or neuropeptide antagonists reduced infarct size and serum norepinephrine level. These findings demonstrate a functional role of spinal astrocytes in myocardial I/R injury and provide a novel potential therapeutic approach targeting spinal cord astrocytes for the prevention of cardiac injury.

Cardiac-targeted PIASy gene silencing mediates deSUMOylation of caveolin-3 and prevents ischemia/reperfusion-induced Nav1.5 downregulation and ventricular arrhythmias

BackgroundAbnormal myocardial Nav1.5 expression and function cause lethal ventricular arrhythmias during myocardial ischemia–reperfusion (I/R). Protein inhibitor of activated STAT Y (PIASy)-mediated caveolin-3 (Cav-3) SUMO modification affects Cav-3 binding to the voltage-gated sodium channel 1.5 (Nav1.5). PIASy activity is increased after myocardial I/R, but it is unclear whether this is attributable to plasma membrane Nav1.5 downregulation and ventricular arrhythmias.MethodsUsing recombinant adeno-associated virus subtype 9 (AAV9), rat cardiac PIASy was silenced using intraventricular injection of PIASy short hairpin RNA (shRNA). After two weeks, rat hearts were subjected to I/R and electrocardiography was performed to assess malignant arrhythmias. Tissues from peri-infarct areas of the left ventricle were collected for molecular biological measurements.ResultsPIASy was upregulated by I/R (P < 0.01), with increased SUMO2/3 modification of Cav-3 and reduced membrane Nav1.5 density (P < 0.01). AAV9-PIASy shRNA intraventricular injection into the rat heart downregulated PIASy after I/R, at both mRNA and protein levels (P < 0.05 vs. Scramble-shRNA + I/R group), decreased SUMO-modified Cav-3 levels, enhanced Cav-3 binding to Nav1.5, and prevented I/R-induced decrease of Nav1.5 and Cav-3 co-localization in the intercalated disc and lateral membrane. PIASy silencing in rat hearts reduced I/R-induced fatal arrhythmias, which was reflected by a modest decrease in the duration of ventricular fibrillation (VF; P < 0.05 vs. Scramble-shRNA + I/R group) and a significantly reduced arrhythmia score (P < 0.01 vs. Scramble-shRNA + I/R group). The anti-arrhythmic effects of PIASy silencing were also evidenced by decreased episodes of ventricular tachycardia (VT), sustained VT and VF, especially at the time 5–10 min after ischemia (P < 0.05 vs. Scramble-shRNA + IR group). Using in vitro human embryonic kidney 293 T (HEK293T) cells and isolated adult rat cardiomyocyte models exposed to hypoxia/reoxygenation (H/R), we confirmed that increased PIASy promoted Cav-3 modification by SUMO2/3 and Nav1.5/Cav-3 dissociation after H/R. Mutation of SUMO consensus lysine sites in Cav-3 (K38R or K144R) altered the membrane expression levels of Nav1.5 and Cav-3 before and after H/R in HEK293T cells.ConclusionsI/R-induced cardiac PIASy activation increased Cav-3 SUMOylation by SUMO2/3 and dysregulated Nav1.5-related ventricular arrhythmias. Cardiac-targeted PIASy silencing mediated Cav-3 deSUMOylation and partially prevented I/R-induced Nav1.5 downregulation in the plasma membrane of cardiomyocytes, and subsequent ventricular arrhythmias in rats. PIASy was identified as a potential therapeutic target for life-threatening arrhythmias in patients with ischemic heart diseases.

Analysis of Anti-Arrhythmic Impacts of Crocin through Estimation of Expression of Cx43 in Myocardial Infarction Using a Rat Animal Model.

Arrhythmia is an important cause of death after myocardial infarction (MI). Different substances have been evaluated for their anti-arrhythmic effect in MI. This study was performed to evaluate the anti-arrhythmic impacts of crocin in an MI animal model (rat) by estimation of the expression of connexin 43 (Cx43). Fifty male Sprague–Dawley rats were grouped into 5 groups, each composed of 10 rats. The first group was regarded as the normal control group and the second one was considered as the MI group, which was caused by ligation of the left anterior descending artery. The other three groups received crocin 50 or 10 mg/kg/day or metoprolol 100 mg/kg/day for 1 week, following ligation of the left anterior descending artery. Evaluated outcomes were cardiac Cx43 expression, arrhythmia incidence, histological findings, and myocyte resting potential. Crocin-treated MI groups showed a significantly lower arrhythmia score than the non-treated MI group, 10 mg/kg/day (1.85 ± 0.55, p < 0.01) and 50 mg/kg/day (1.70 ± 0.33, p < 0.01). Groups that received crocin 10 mg/kg/day (66.30 ± 2.59, p < 0.01), crocin 50 mg/kg/day (68.10 ± 2.43, p < 0.01), and metoprolol 100 mg/kg/day (−63.54 ± 0.63 mV, p < 0.01) significantly prevented depolarization in comparison with the non-treated MI group. Expression of Cx43 mRNA in crocin 10 mg/kg/day (1.54 ± 0.24, p < 0.01), crocin 50 mg/kg/day (1.73 ± 0.09, p < 0.01), and metoprolol 100 mg/kg/day (1.75 ± 0.14, p < 0.01) treatment groups was significantly higher in comparison with the non-treated MI group. Crocin showed a preventive effect on the arrhythmogenic impact of MI in an experimental model of ischemic injury through an increase in expression of Cx43.

Myeloid differentiation factor 2 inhibitor and N-acetyl cysteine synergistically reduced left ventricular dysfunction in rats with cardiac ischemia/reperfusion injury

Abstract Background Myeloid differentiation factor 2 inhibitor (MD2i) is a novel anti-inflammatory agent that exerts favorable outcomes in various diseases including cardiac ischemia/reperfusion (I/R) injury. However, whether a potent antioxidant N-acetyl cysteine (NAC) can augment the beneficial effects of MD2i in rats with cardiac I/R injury have never been investigated. Purpose We tested the hypothesis that NAC increases the beneficial effects of MD2i against cardiac I/R injury in rats. Methods Rats were divided into either a sham (n=6) or cardiac I/R group (n=72). Rats in the I/R group received one of the following 6 treatments (n=12 each) at the onset of reperfusion: vehicle, MD2i at 20 and 40 mg/kg, NAC at 75 and 150 mg/kg, and combined MD2i 20 mg/kg with NAC 150 mg/kg. Left ventricular (LV) function, infarct size, arrhythmia score, and cardiac mitochondrial function and dynamics were determined. Results Myocardial infarction, LV dysfunction, and cardiac arrhythmias were observed in rats with cardiac I/R injury, along with mitochondrial dysfunction (Fig. 1). Treatment with MD2i at either 20 or 40 mg/kg effectively reduced LV dysfunction but failed to reduce the infarct size (Fig. 1). NAC at 150 mg/kg, but not at 75 mg/kg, significantly decreased both LV dysfunction and infarct size following cardiac I/R injury (Fig. 1). However, combined treatment exerted even greater efficacy in reducing cardiac I/R injury than monotherapy, through a greater reduction of cardiac mitochondrial dysfunction and mitochondrial fission (Fig. 1). However, no benefit on reducing the arrhythmia score in all groups. Conclusion Combined MD2i and NAC treatment exerted a superior cardioprotective effect against cardiac I/R injury than either monotherapy regimen via an improved cardiac mitochondrial function. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): 1. NSTDA Research Chair Grant from the National Science and Technology Development Agency Thailand2. National Research Council of Thailand

Effects of Glucose Fluctuations on Electrocardiogram Readings and the Development of Ventricular Arrhythmia in Diabetic Rats

Objective: Many studies have shown that blood glucose fluctuations (GFs) are more harmful to patients with diabetes mellitus (DM) than sustained hyperglycemia. However, the effect of GF on electrocardiogram (ECG) parameters and vulnerability to ventricular tachycardia/fibrillation (VT/VF) remains poorly characterized. This study aimed to assess the effect of GF on ECG parameters and induction of VT/VF in streptozotocin (STZ)-induced diabetic Sprague-Dawley rats. Methods: Male Sprague-Dawley rats were injected with STZ and randomly divided into 3 treatment groups: controlled STZ-induced diabetic rats (C-STZ) (n = 10); uncontrolled STZ-induced diabetic rats (U-STZ) (n = 10); and STZ-induced diabetic rats with glucose fluctuations (GF-STZ) (n = 10). After 12 weeks, baseline ECG recordings were taken and a VT/VF test was performed with the administration of caffeine and dobutamine. Hematoxylin &amp; eosin and masson staining were used to evaluate pathological cardiac changes after intervention. Results: No significant difference in heart rate, RR interval, P wave (duration and height), PR segment, PR interval, QRS wave duration, and T wave height was observed among the 3 groups (P &gt; 0.05). Compared with the C-STZ group, the U-STZ and GF-STZ groups both had a longer T wave duration ((62.41 ± 2.38) ms vs. (78.37 ± 4.64) ms and (96.06 ± 4.60) ms, P &lt; 0.05), QT interval ((83.66 ± 2.31) ms vs. (101.75 ± 4.56) ms and (119.14 ± 4.88) ms, P &lt; 0.05), and QTc interval ((77.45 ± 1.36) ms vs. (91.36 ± 3.49) ms and (104.55 ± 3.01) ms, P &lt; 0.05), all of which were longest in the GF-STZ group (P &lt; 0.05). Additionally, the GF-STZ group had the highest VT/VF occurrence and duration and the highest arrhythmia score. Conclusion: This study revealed GF can significantly prolong the QT interval, QTc interval, and T wave duration, as well as increase vulnerability to VT/VF in rats, which may be an important electrophysiological mechanism of GF-related ventricular arrhythmia.

Effect of rostral ventrolateral medulla in electroacupuncture pretreatment for alleviating myocardial ischemia-reperfusion injury 延髓头端腹外侧区在电针预处理减轻心肌缺血再灌注损伤中的作用

ObjectiveTo observe the effects of pretreatment with electroacupuncture (EA) on neuron activity in the rostral ventrolateral medulla (RVLM) of rats with myocardial ischemia-reperfusion injury (MIRI) and explore the central regulatory mechanism of EA in attenuating MIRI. MethodsOf 72 SD rats, 12 were randomly allocated into the group of EA pretreatment + RVLM nucleus damage (EA + RVLM lesion group). The other 60 rats were randomized (20 rats each) into a sham-operation group, a model group, and an EA pretreatment group (EA group). Except for the rats in the sham-operation group, the models of MIRI were prepared by ligating the left anterior descending coronary artery in the model, EA, and EA + RVLM lesion groups. The rats of the EA group were intervened with EA at “Shénmén (神门HT7)” and “Tōnglĭ (通里HT5), 1 mA in current intensity and 2 Hz in frequency, for 20 min each time per day. Before modeling, the intervention was given for seven consecutive days. In the EA + RVLM lesion group, 3 weeks after microinjection with the neuronal apoptotic virus at bilateral RVLM, the same EA intervention as the EA group was provided. Afterward, the MIRI models were prepared. In the model group, no EA intervention was given. Using Powerlab electrophysiolograph, ST segment displacement value and arrhythmia score were recorded and analyzed before modeling, 30 min after ligation, and 120 min after reperfusion in each group. The concentration of cardiac troponin (cTnl) was detected with an ELISA assay kit. Using immunofluorescence staining, the expression level of c-fos protein of RVLM was detected in the sham-operation, model, and EA groups separately. Plexon multichannel acquisition processor was adopted to record the neuronal firing and field potential of RVLM in the sham-operation, model, and EA groups. ResultsST segment displacement value, arrhythmia score, and cTnl concentration 30 min after ligation and 120 min after reperfusion were all elevated in the model group compared to the sham-operation group (all P < 0.01). ST segment displacement value, arrhythmia score, and cTnl concentration were lower in the EA group compared to the model and EA + RVLM lesion groups (P < 0.01). Compared with the sham-operation group, the expression level of c-fos and the total firing frequency in RVLM were significantly higher in the model group (P < 0.01). However, the expression level of c-fos and the total firing frequency in RVLM were lower in the EA group compared with the model group (P < 0.01). ConclusionElectroacupuncture pretreatment may induce changes in c-fos protein expression and neuronal activity in RVLM to mitigate myocardial lesions. RVLM plays an important role in electroacupuncture pretreatment for alleviating MIRI.

Rosiglitazone Does Not Show Major Hidden Cardiotoxicity in Models of Ischemia/Reperfusion but Abolishes Ischemic Preconditioning-Induced Antiarrhythmic Effects in Rats In Vivo.

Clinical observations are highly inconsistent with the use of the antidiabetic rosiglitazone regarding its associated increased risk of myocardial infarction. This may be due to its hidden cardiotoxic properties that have only become evident during post-marketing studies. Therefore, we aimed to investigate the hidden cardiotoxicity of rosiglitazone in ischemia/reperfusion (I/R) injury models. Rats were treated orally with either 0.8 mg/kg/day rosiglitazone or vehicle for 28 days and subjected to I/R with or without cardioprotective ischemic preconditioning (IPC). Rosiglitazone did not affect mortality, arrhythmia score, or infarct size during I/R. However, rosiglitazone abolished the antiarrhythmic effects of IPC. To investigate the direct effect of rosiglitazone on cardiomyocytes, we utilized adult rat cardiomyocytes (ARCMs), AC16, and differentiated AC16 (diffAC16) human cardiac cell lines. These were subjected to simulated I/R in the presence of rosiglitazone. Rosiglitazone improved cell survival of ARCMs at 0.3 μM. At 0.1 and 0.3 μM, rosiglitazone improved cell survival of AC16s but not that of diffAC16s. This is the first demonstration that chronic administration of rosiglitazone does not result in major hidden cardiotoxic effects in myocardial I/R injury models. However, the inhibition of the antiarrhythmic effects of IPC may have some clinical relevance that needs to be further explored.

IKs inhibitor JNJ303 prolongs the QT interval and perpetuates arrhythmia when combined with enhanced inotropy in the CAVB dog

IntroductionImpaired IKs induced by drugs or due to a KCNQ1 mutation, diagnosed as long QT syndrome type 1 (LQT1) prolongs the QT interval and predisposes the heart to Torsade de Pointes (TdP) arrhythmias. The anesthetized chronic AV block (CAVB) dog is inducible for TdP after remodeling and IKr inhibitor dofetilide. We tested the proarrhythmic effect of IKs inhibition in the CAVB dog, and the proarrhythmic role of increased contractility herein. MethodsDofetilide-inducible animals were included to test the proarrhythmic effect of 1) IKs inhibition by JNJ303 (0.63 mg/kg/10min i.v.; n = 4), 2) IKs inhibition combined with enhanced inotropy (ouabain, 0.045 mg/kg/1min i.v.; n = 6), and 3) the washout period of the anesthetic regime (n = 10). ResultsJNJ303 prolonged the QTc interval (from 477 ± 53 ms to 565 ± 14 ms, P < 0.02) resembling standardized dofetilide-induced QTc prolongation. Single ectopic beats (n = 4) and ventricular tachycardia (VT) (n = 3) were present, increasing the arrhythmia score (AS) from 1.0 ± 0 to 7.1 ± 6.5. JNJ303 combined with ouabain increased contractile parameters (LVdP/dtmax from 1725 ± 273 to 4147 ± 611 mmHg/s, P < 0.01). Moreover, TdP arrhythmias were induced in 4/6 dogs and AS increased from 1.0 ± 0 to 20.2 ± 19.0 after JNJ303 and ouabain (P < 0.05). Finally, TdP arrhythmias were induced in 4/10 dogs during the anesthesia washout period and the AS increased from 1.1 ± 0.3 to 9.2 ± 11.2. ConclusionMimicking LQT1 using IKs inhibitor JNJ303 prolongs the QTc interval and triggers ectopic beats and non-sustained VT in the CAVB dog. Induction of the more severe arrhythmic events (TdP) demands a combination of IKs inhibition with enhanced inotropy or ending the anesthetic regime.

(-)-Carvone Modulates Intracellular Calcium Signaling with Antiarrhythmic Action in Rat Hearts.

ResumoFundamento:A (-)-carvona é um monoterpeno encontrado em óleos essenciais com atividade antioxidante e anti-inflamátoria.Objetivos:O objetivo deste estudo foi analisar a propriedade antiarrítmica da (-)-carvona no coração de rato e seus efeitos sobre a sinalização de Ca+2 intracelular.Métodos:Os efeitos da (-)-carvona foram avaliados sobre a contratilidade atrial (0,01 – 4 mM) e ventricular (0,5 mM), e no eletrocardiograma (0,5mM). A fração de encurtamento, a corrente de cálcio do tipo L (ICa,L) e a sinalização de Ca+2 foram medidas no cardiomiócito isolado (0,5 mM). O efeito antiarrítmico foi avaliado no modelo de arritmia induzida por sobrecarga de cálcio (0,5 mM) (n = 5). Um p < 0,05 foi adotado como nível de significância estatística.Resultados:No átrio, a (-)-carvona causou inotropismo negativo de maneira concentração-dependente (EC50 0,44 ± 0,11 mM) e diminuiu o inotropismo positivo induzido pelo CaCl2 (0,1 – 8,0 mM) e BAY K8644 (5 - 500 nM), um agonista de canal de cálcio do tipo L. Em coração isolado, a (-)-carvona (0,5mM) reduziu a contratilidade ventricular em 73% e a frequência cardíaca (em 46%), aumentou o Pri (30,7%, tempo desde o início da onda P até a onda R) e o QTc (9,2%, uma medida de despolarização e repolarização dos ventrículos), sem mudar a duração do complexo QRS. A (-)-carvona diminuiu a fração de encurtamento (61%), a (ICa,L) (79%) e o transiente intracelular de Ca+2 (38%). Além disso, a (-)-carvona apresentou ação antiarrítmica, identificada pela redução do escore de arritmia (85%) e ocorrência de fibrilação ventricular.Conclusão:A (-)-carvona reduz a entrada de Ca+2 através de canais de Ca+2 do tipo L e, assim, diminui a contratilidade cardíaca e o Ca+2 intracelular e apresenta promissora atividade antiarrítmica no coração de ratos.

The temporal impact of erythropoietin administration on mitochondrial function and dynamics in cardiac ischemia/reperfusion injury

Erythropoietin (EPO) has been shown to provide protection against ischemia/reperfusion (I/R) injury in various experimental models. However, clinical trials revealed unsatisfactory results when EPO was given to patients with myocardial infarction following reperfusion. The timing of EPO administration and its relation to mitochondrial function may largely involve in this controversy. We hypothesized that EPO given at different time points exert varying cardioprotective effects in terms of myocardial infarct size, left ventricular (LV) function, arrhythmia, apoptosis, mitochondrial function, and mitochondrial dynamics in rats with cardiac I/R injury. Male Wistar rats were subjected to either sham (n = 6) or cardiac I/R operation (n = 48). Rats undergoing cardiac I/R operation (30-min ischemia, followed by 120-min reperfusion) were allotted into 4 subgroups (n = 12/group): vehicle, EPO pretreatment, EPO given during ischemia, and EPO given at reperfusion. EPO was administered intravenously at 5000 unit/kg. Arrhythmia and LV function were monitored throughout the protocol. Next, the hearts were collected to determine infarct size, mitochondrial function, mitochondrial dynamics, gap junction protein, and apoptosis. Cardiac I/R promoted arrhythmias, LV dysfunction, infarct size expansion, apoptosis, mitochondrial dysfunction and increased mitochondrial fission. EPO given either before or during ischemia, but not at reperfusion, attenuated arrhythmia scores, LV dysfunction, infarct size, and apoptosis. Only EPO given either before or during ischemia alleviated mitochondrial swelling, mitochondrial depolarization, and reduced the levels of p-Drp1ser616/Drp1. Data from in vitro study also confirmed that EPO directly attenuated mitochondrial dysfunction in H9c2 cells subjected to hypoxia/reoxygenation. In conclusion, the EPO administration, either before or during ischemia, exerted cardioprotection against I/R injury by attenuating mitochondrial dynamic imbalance, mitochondrial dysfunction, and apoptosis, leading to reduced infarct size and improved LV function.

Cardioprotective effects of sinomenine in myocardial ischemia/reperfusion injury in a rat model

BackgroundIschemia reperfusion (I/R) play an imperative role in the expansion of cardiovascular disease. Sinomenine (SM) has been exhibited to possess antioxidant, anticancer, anti-inflammatory, antiviral and anticarcinogenic properties. The aim of the study was scrutinized the cardioprotective effect of SM against I/R injury in rat. MethodsRat were randomly divided into normal control (NC), I/R control and I/R + SM (5, 10 and 20 mg/kg), respectively. Ventricular arrhythmias, body weight and heart weight were estimated. Antioxidant, inflammatory cytokines, inflammatory mediators and plasmin system indicator were accessed. ResultsPre-treated SM group rats exhibited the reduction in the duration and incidence of ventricular fibrillation, ventricular ectopic beat (VEB) and ventricular tachycardia along with suppression of arrhythmia score during the ischemia (30 and 120 min). SM treated rats significantly (P < 0.001) altered the level of antioxidant parameters. SM treatment significantly (P < 0.001) repressed the level of creatine kinase MB (CK-MB), creatine kinase (CK) and troponin I (Tnl). SM treated rats significantly (P < 0.001) repressed the tissue factor (TF), thromboxane B2 (TXB2), plasminogen activator inhibitor 1 (PAI-1) and plasma fibrinogen (Fbg) and inflammatory cytokines and inflammatory mediators. ConclusionOur result clearly indicated that SM plays anti-arrhythmia effect in I/R injury in the rats via alteration of oxidative stress and inflammatory reaction.

Preparation, physicochemical characterization, docking and antiarrhythmic effect of d-limonene and d-limonene hydroxypropyl-β-cyclodextrin complex

d-limonene (D-LIM) is a monoterpene found mainly in the essential oil of citrus fruits. Cardiovascular effects of D-LIM alone have been extensively demonstrated. Despite possessing important pharmacological effects, its chemical properties prevent its clinical application. This study aimed to produce and characterize hydroxypropyl-β-cyclodextrin (HP-β-CD) inclusion complex with D-LIM, and to assess its cardiovascular effects in animal model of arrhythmia. The inclusion complex was prepared by the slurry complexation, and characterized using HPLC, PXRD, DSC, and 1H NMR. Molecular docking was performed to analyze the interactions between the complex. The antiarrhythmic effect was assessed in an animal model of arrhythmia induced by Bay K 8644. The physicochemical characterization showed that the D-LIM + HP-β-CD complex formation had a complexation efficiency of 79.96 ± 0.24%, which was corroborated with all analysis performed. Docking showed that D-LIM interacts with HP-β-CD through hydrophobic and van der Waals bonds, with binding affinity of −4.2 kcal/mol. D-LIM + HP-β-CD complex (10 mg/kg, i.v.) significantly reduced arrhythmias, reducing arrhythmia score (p < 0.01), duration of arrhythmias (p < 0.05) and prevented tachycardia (p < 0.05). Our results showed an efficient method of complexation of D-LIM + HP-β-CD and improved its cardiovascular effects compared to D-LIM.

Effect of TLR4/MyD88/NF-kB axis in paraventricular nucleus on ventricular arrhythmias induced by sympathetic hyperexcitation in post-myocardial infarction rats.

Sympathetic activation after myocardial infarction (MI) leads to ventricular arrhythmias (VAs), which can result in sudden cardiac death (SCD). The toll‐like receptor 4 (TLR4)/myeloid differentiation primary response 88 (MyD88)/nuclear factor‐kappa B (NF‐kB) axis within the hypothalamic paraventricular nucleus (PVN), a cardiac‐neural sympathetic nerve centre, plays an important role in causing VAs. An MI rat model and a PVN‐TLR4 knockdown model were constructed. The levels of protein were detected by Western blotting and immunofluorescence, and localizations were visualized by multiple immunofluorescence staining. Central and peripheral sympathetic activation was visualized by immunohistochemistry for c‐fos protein, renal sympathetic nerve activity (RSNA) measurement, heart rate variability (HRV) analysis and norepinephrine (NE) level detection in serum and myocardial tissue measured by ELISA. The arrhythmia scores were measured by programmed electrical stimulation (PES), and cardiac function was detected by the pressure–volume loop (P‐V loop). The levels of TLR4 and MyD88 and the nuclear translocation of NF‐kB within the PVN were increased after MI, while sympathetic activation and arrhythmia scores were increased and cardiac function was decreased. However, inhibition of TLR4 significantly reversed these conditions. PVN‐mediated sympathetic activation via the TLR4/MyD88/NF‐kB axis ultimately leads to the development of VAs after MI.

Acacetin alleviates myocardial ischaemia/reperfusion injury by inhibiting oxidative stress and apoptosis via the Nrf-2/HO-1 pathway

Context Acacetin is a natural source of flavonoids with anti-inflammatory and antioxidant effects. Objective This study determines acacetin’s protective effect and mechanism on myocardial ischaemia/reperfusion (I/R) injury. Materials and methods Sprague-Dawley rats were divided into sham and I/R injury and treatment with acacetin. Acacetin (10 mg/kg) was subcutaneously injected for 7 days. ECG and echocardiography were conducted to determine arrhythmia and heart function. The pathological characters of the heart were determined with triphenyl tetrazolium chloride staining, Haematoxylin & Eosin staining, and Masson staining. Expression of proteins in infarct tissues was examined with western blots. Results Administrated with acacetin in I/R rats significantly reduced the arrhythmia score from 4.90 to 2.50 and the reperfusion arrhythmia score from 3.79 to 1.82 in the vehicle or the acacetin group, respectively. LVEF was improved from 33.5% in the I/R group to 43.7% in the acacetin group, LVFS was increased from 16.4% to 24.5%, LVIDs was decreased from 6.5 to 5.3 mm. The inflammatory cell infiltration, myocardial fibrosis, and collagen 1 and 3 were reduced by acacetin. Acacetin promoted SOD and decreased MDA. In myocardial tissues, the expression level of TLR4 and IL-6 were restrained, and IL-10 was promoted. Apoptotic protein Bax was suppressed, and anti-apoptotic protein Bcl-2 was promoted in the acacetin group. Interestingly, the transcription factor Nrf-2/HO-1 pathway was also reversed by acacetin. Discussion and conclusion Our findings indicated that acacetin has a potential therapeutic effect in clinical application on treating I/R-induced heart injury.